Epidermal growth factor alters silica nanoparticle uptake and improves gold-nanoparticle-mediated gene silencing in A549 cells

Introduction: Delivery of therapeutic nanoparticles (NPs) to cancer cells represents a promising approach for biomedical applications. A key challenge for nanotechnology translation from the bench to the bedside is the low amount of administered NPs dose that effectively enters target cells. To improve NPs delivery, several studies proposed NPs conjugation with ligands, which specifically deliver NPs to target cells via receptor binding. One such example is epidermal growth factor (EGF), a peptide involved in cell signaling pathways that control cell division by binding to epidermal growth factor receptor (EGFR). However, very few studies assessed the influence of EGF present in the cell environment, on the cellular uptake of NPs.Methods: We tested if the stimulation of EGFR-expressing lung carcinomacells A549 with EGF affects the uptake of 59 nm and 422 nm silica (SiO2) NPs. Additionally, we investigated whether the uptake enhancement can be achieved with gold NPs, suitable to downregulate the expression of cancer oncogene c-MYC.Results: Our findings show that EGF binding to its receptor results in receptor autophosphorylation and initiate signaling pathways, leading to enhanced endocytosis of 59 nm SiO2 NPs, but not 422 nm SiO2 NPs. Additionally, we demonstrated an enhanced gold (Au) NPs endocytosis and subsequently a higher downregulation of c-MYC.Discussion: These findings contribute to a better understanding of NPs uptake in the presence of EGF and that is a promising approach for improved NPs delivery

Intralinguale Untertitelung aus variationslinguistischer Sicht am Beispiel von "Komm, süßer Tod" und "Hinterholz 8"

In vorliegender Diplomarbeit geht es um die Untersuchung zweier österreichischer Filme im Hinblick auf deren standarddeutsche Untertitel, die im zweiten Teil meiner Arbeit, dem empirischen Teil, vorgestellt wird. Um in dieses Thema einzuführen, dient der vorangestellte theoretische Teil bestehend aus drei Kapiteln. Die Arbeit behandelt folgende Forschungsfragen: Welche Funktion hat die Untertitelung in ein- und dieselbe Sprache und ist diese stringent? Welche Funktion hat der Nonstandard in den von mir behandelten Filmen

Quantitative nuclear feature analysis in the prognosis of benign breast disease and ductal carcinoma in situ

Except of the diagnostic categories based on the morphology of cells and tissue there are currently no significant prognostic markers for patients with benign breast disease or ductal carcinoma in situ (DCIS). This thesis proposed that such prognostic information could be obtained based on quantitative nuclear feature analysis of diagnostic cells and/or normal appearing cells in breast tissue. Image cytometry (IC) measurements provide numerous quantitative nuclear features which reflect the DNA content, nuclear morphology and chromatin condensation patterns in stained nuclei. With the use of this technique it is possible to detect slight morphological nuclear changes which can not be observed by the human eye. Measurements on tissue sections were chosen for this study because in these specimens visualization of tissue histology and precise identification of affected glands is possible. The thesis involved testing of the following postulates: 1) DNA ploidy can be evaluated by IC measurements of tissue sections DNA content measurements of archival breast specimens showed that the performance of manual IC measurements of nuclei on tissue sections was comparable to the results of flow cytometry and automated IC techniques. 2) Quantitative nuclear features change in different histological patterns of breast diseases The analysis of normal tissue, non-proliferative breast disease, proliferative breast disease, carcinoma in situ and invasive cancer specimens demonstrated differences in nuclear features between different h istopatholog ical patterns. Compared to normal tissue characteristics, the deviations of features related to the nuclear area, shape, DNA content and chromatin texture all increased with advancing morphological changes. 3) Various histological types of DCIS can be characterized on the basis of the nuclear features The differences in quantitative nuclear features were defined for various histological types of DCIS. The nuclear DNA content, size, irregularity of shape and chromatin texture, all increased from the lowest values in cribriform type to the highest values in comedo type DCIS. Aneuploidy was demonstrated in about 60% of non-comedo DCIS and in about 95% of comedo DCIS. 4) Differences in nuclear morphology, which may be related to the invasive potential of ductal carcinomas in situ (DCIS), exist between I) pure DCIS, and ii) DCIS with synchronous invasive carcinoma An important task of this thesis was to obtain a prognostic indicator for ductal carcinoma in situ (DCIS). Nuclear features of DCIS which were associated with the presence of invasive carcinoma in the surrounding breast tissue were identified. A classification system based on these nuclear features was then used to discriminate between cases with pure DCIS and cases of DCIS with invasive cancer in the surrounding tissue. This classification predicted accurately the presence of invasive carcinoma in about 80% of non-comedo DCIS and in about 100% of comedo DCIS cases. 5) Subtle changes of nuclear morphology exist in epithelial cells of normal appearing breast tissue adjacent to invasive carcinoma (malignancy associated changes) The final goal was to test the hypothesis that small deviations in nuclear morphology, indicative of the malignancy in the surrounding tissue, can be detected through nuclear measurements of non-diagnostic, normal appearing cells in the vicinity of DCIS or invasive carcinoma. This phenomenon has been previously studied in other tissues and has been described as a part of malignancy associated changes (MAC). The present study illustrated the existence of MAC in the normal appearing breast tissue adjacent to in situ, or invasive carcinoma. Patients with benign or malignant diseases could be distinguished solely on the basis of the measurements of epithelial nuclei from the normal appearing glands. With the analysis of MAC about 85% of cases were correctly classified as benign or malignant. In conclusion: A) Differences in nuclear features which were demonstrated for various groups of breast diseases suggest that these features could be applied as an objective aid in the classification and diagnosis of breast diseases. B) The analysis of DCIS nuclei can provide useful prognostic information making it possible to suspect the presence of invasive carcinoma in the breast when only DCIS is present in the biopsy. Moreover, specific changes in nuclear morphology, which are characteristic of DCIS associated with invasive carcinoma in the surrounding breast might be associated with the progressive capacity of DCIS and may be helpful as a marker predictive of the subsequent behavior of DIS tumors. C) Nuclear features, characteristic of MAC, are important as markers for occult malignancy in cases where only benign breast disease is found in a biopsy. In addition, a high frequency of “MAC” nuclei in a benign breast tissue may be suggestive of a higher progressive potential and of an increased risk for the development of invasive carcinoma from benign breast tissue which has a high frequency of “MAC” nuclei. In view of the clinical relevance of these findings it is very important to confirm and expand the results with further studies on larger number of patients.Medicine, Faculty ofPathology and Laboratory Medicine, Department ofGraduat

Current Perspectives on Lobular Neoplasia of the Breast

Purpose of Review To present current data on histopathologic types, imaging presentation, and management of lobular neoplasia of the breast. Recent Findings Non-invasive lobular neoplasia is a heterogeneous group of lesions, including classic lobular neoplasia, composed of atypical lobular neoplasia and classic lobular carcinoma in situ, and lobular neoplasia variants, composed of pleomorphic lobular carcinoma in situ and florid lobular carcinoma in situ. The incidence of lobular neoplasia as the primary diagnosis on percutaneous core needle biopsies ranges from 1 to 2%. Classic lobular neoplasia is typically an incidental finding, while lobular neoplasia variants are typically the imaging target and, like ductal carcinoma in situ, most commonly present as calcifications. Current recommendations for classic lobular neoplasia on core needle biopsy favor nonsurgical approaches, including observation and chemoprevention. Surgical excisional biopsy is warranted if biopsy demonstrates pleomorphic and/or florid lobular carcinoma in situ, due to the high prevalence of an associated invasive carcinoma. Excision is also required for classic lobular neoplasia with discordant radiologic-pathologic findings. Additionally, lobular neoplasia is a risk factor for future development of breast cancer, and these patients require consultation for risk assessment and counseling for surveillance and risk reduction. Summary This review presents current histopathologic types of lobular neoplasia, their respective risks, and management strategies. Following a diagnosis on core needle biopsy, a multispecialty team approach is essential for adequate management

Low to Intermediate (Borderline) Grade Breast Spindle Cell Lesions on Needle Biopsy: Diagnostic Approach and Clinical Management

Spindle cell proliferations of the breast are a heterogeneous group of lesions ranging from benign or reactive lesions to aggressive malignant neoplasms. Diagnosis on core biopsy can be particularly challenging as lesions displaying different lineages associated with variable outcomes share overlapping morphologies (scar vs. fibromatosis-like metaplastic carcinoma) whereas individual entities can exhibit a large variety of appearances (myofibroblastoma). In this review, lesions are grouped into lineage, when possible, including those showing fibroblastic/myofibroblastic differentiation, ranging from entities that require no additional management, such as scar and nodular fasciitis, to those with unpredictable clinical outcomes such as fibromatosis and solitary fibrous tumor or locally aggressive behavior such as dermatofibrosarcoma protuberans. The review of low-grade vascular lesions includes atypical vascular lesion and low-grade angiosarcoma. Also discussed are various adipocytic lesions ranging from lipoma to liposarcoma, and rare smooth muscle and neural entities more commonly encountered in locations outside the breast, such as leiomyoma, neurofibroma, schwannoma, or granular cell tumor. Optimal histological evaluation of these entities merges clinical and radiologic data with morphology and ancillary testing. We present our approach to immunohistochemical and other ancillary testing and highlight issues in pathology correlation with imaging. Recent updates in the management of breast spindle cell lesions are addressed. In a well-sampled lesion with radiographic concordance, the core biopsy diagnosis reliably guides management and we advocate the inclusion of management recommendations in the pathology report. Precise characterization using up to date guidelines is important to identify a subset of patients who may safely avoid unnecessary surgical procedures. A multidisciplinary approach with close collaboration with our clinical colleagues is emphasized

Increased uptake of silica nanoparticles in inflamed macrophages but not upon co-exposure to micron-sized particles

Silica nanoparticles (NPs) are widely used in various industrial and biomedical applications. Little is known about the cellular uptake of co-exposed silica particles, as can be expected in our daily life. In addition, an inflamed microenvironment might affect a NP’s uptake and a cell’s physiological response. Herein, prestimulated mouse J774A.1 macrophages with bacterial lipopolysaccharide were post-exposed to micron- and nanosized silica particles, either alone or together, i.e., simultaneously or sequentially, for different time points. The results indicated a morphological change and increased expression of tumor necrosis factor alpha in lipopolysaccharide prestimulated cells, suggesting a M1-polarization phenotype. Confocal laser scanning microscopy revealed the intracellular accumulation and uptake of both particle types for all exposure conditions. A flow cytometry analysis showed an increased particle uptake in lipopolysaccharide prestimulated macrophages. However, no differences were observed in particle uptakes between single- and co-exposure conditions. We did not observe any colocalization between the two silica (SiO2) particles. However, there was a positive colocalization between lysosomes and nanosized silica but only a few colocalized events with micro-sized silica particles. This suggests differential intracellular localizations of silica particles in macrophages and a possible activation of distinct endocytic pathways. The results demonstrate that the cellular uptake of NPs is modulated in inflamed macrophages but not in the presence of micron-sized particles

The Functions of Cholera Toxin Subunit B as a Modulator of Silica Nanoparticle Endocytosis

The gastrointestinal tract is the main target of orally ingested nanoparticles (NPs) and at the same time is exposed to noxious substances, such as bacterial components. We investigated the interaction of 59 nm silica (SiO2) NPs with differentiated Caco-2 intestinal epithelial cells in the presence of cholera toxin subunit B (CTxB) and compared the effects to J774A.1 macrophages. CTxB can affect cellular functions and modulate endocytosis via binding to the monosialoganglioside (GM1) receptor, expressed on both cell lines. After stimulating macrophages with CTxB, we observed notable changes in the membrane structure but not in Caco-2 cells, and no secretion of the pro-inflammatory cytokine tumor necrosis factor-α (TNF-α) was detected. Cells were then exposed to 59 nm SiO2 NPs and CtxB sequentially and simultaneously, resulting in a high NP uptake in J774A.1 cells, but no uptake in Caco-2 cells was detected. Flow cytometry analysis revealed that the exposure of J774A.1 cells to CTxB resulted in a significant reduction in the uptake of SiO2 NPs. In contrast, the uptake of NPs by highly selective Caco-2 cells remained unaffected following CTxB exposure. Based on colocalization studies, CTxB and NPs might enter cells via shared endocytic pathways, followed by their sorting into different intracellular compartments. Our findings provide new insights into CTxB’s function of modulating SiO2 NP uptake in phagocytic but not in differentiated intestine cells

Interleukin 17 receptor A modulates monocyte subsets and macrophage generation in vivo.

Interleukin (IL)-17A signaling via Interleukin 17 receptor A (Il17ra) contributes to the inflammatory host response by inducing recruitment of innate immune cells, but also plays a role in homeostatic neutrophilic granulocyte regulation. Monocytes, the other main innate immune cell, have a longer life span and can pursue multiple differentiation pathways towards tissue macrophages. Monocytes are divided into two subpopulations by expression of the Ly6C/Gr1 surface marker in mice. We here investigated the role of Il17ra in monocyte homeostasis and macrophage generation. In Il17ra(-/-) and in mixed bone marrow chimeric wt/Il17ra(-/-) mice, the concentrations of circulating Il17ra(-/-) Gr1(low) monocytes were significantly decreased compared to wt cells. Pulmonary, splenic and resident peritoneal Il17ra(-/-) macrophages were significantly fewer than of wt origin. Bone marrow progenitor and monocyte numbers were equal, but the proportion of Il17ra(-/-) Gr1(low) monocytes was already decreased at bone marrow level. After monocyte depletion, initial Gr1(high) and Gr1(low) monocyte regeneration of Il17ra(-/-) and wt cells was very similar. However, Il17ra(-/-) Gr1(low) counts were not sustained. After labeling with either fluorescent beads or BrdU, Il17ra(-/-) Gr1(high) monocyte transition to Gr1(low) cells was not detectable unlike wt cells. Monocyte recruitment in acute peritonitis, which is known to be largely due to Gr1(high) cell migration, was unaffected in an identical environment. Unilateral ureteral obstruction induces a less acute inflammatory and fibrotic kidney injury. Compared to wt cells in the same environment, Il17ra(-/-) macrophage accumulation in the kidney was decreased. In the absence of Il17ra on all myeloid cells, renal fibrosis was significantly attenuated. Our data show that Il17ra modulates Gr1(low) monocyte counts and suggest defective Gr1(high) to Gr1(low) monocyte transition as an underlying mechanism. Lack of Il17ra altered homeostatic tissue macrophage formation and diminished renal inflammation and fibrosis. Il17ra appears to be a novel modulator of monocyte phenotype and possible therapeutic target in renal fibrosis